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Aquaporin-4 (AQP4) plays a crucial role in brain water circulation and is considered a therapeutic target in hydrocephalus. Congenital hydrocephalus is associated with a reaction of astrocytes in the periventricular white matter both in experimental models and human cases. A previous report showed that bone marrow-derived mesenchymal stem cells (BM-MSCs) transplanted into the lateral ventricles of hyh mice exhibiting severe congenital hydrocephalus are attracted by the periventricular astrocyte reaction, and the cerebral tissue displays recovery. The present investigation aimed to test the effect of BM-MSC treatment on astrocyte reaction formation. BM-MSCs were injected into the lateral ventricles of four-day-old hyh mice, and the periventricular reaction was detected two weeks later. A protein expression analysis of the cerebral tissue differentiated the BM-MSC-treated mice from the controls and revealed effects on neural development. In in vivo and in vitro experiments, BM-MSCs stimulated the generation of periventricular reactive astrocytes overexpressing AQP4 and its regulatory protein kinase D-interacting substrate of 220 kDa (Kidins220). In the cerebral tissue, mRNA overexpression of nerve growth factor (NGF), vascular endothelial growth factor (VEGF), hypoxia-inducible factor-1 (HIF1α), and transforming growth factor beta 1 (TGFβ1) could be related to the regulation of the astrocyte reaction and AQP4 expression. In conclusion, BM-MSC treatment in hydrocephalus can stimulate a key developmental process such as the periventricular astrocyte reaction, where AQP4 overexpression could be implicated in tissue recovery.

[EN] Reducing mechanical losses in internal combustion engines has been a recurrent research topic over the past few decades. Despite mechanical losses are a key issue that should be carefully addressed to reduce fuel consumption and emissions, its distribution amongst engine elements is barely covered in literature. Recent work has shown the potential advantage of using low viscosity engine oils to reduce fuel consumption, however, there is reduced knowledge on mechanical losses distribution under transient conditions. In this work, a model is presented that predicts not only the total friction losses of an engine, but determines the amount of friction energy lost in piston-ring assembly, engine bearings, camshaft and engine auxiliaries in driving cycles representing a real driving route. The stationary mechanical losses engine maps can be used due to non-dependence on mechanical losses with temperature in warm driving cycle conditions. The final results of the simulation predicts, with 2% error, fuel consumption, energy expended by the driven wheel and mechanical losses of the engine. This methodology reduces the computational cost to estimate key engine parameters in a real driving cycle such as: mechanical losses and its distribution, fuel consumption... as the total calculation time is 10 s per cycle simulated., The authors would like to thank different members of the CMT-Motores Termicos team of the Universitat Politecnica de Valencia for their contribution to this work. The authors would also like to thank the Spanish Ministry of Science, Innovation and Universities and Universities for financing the PhD. Studies of Antonio J. Jimenez-Reyes (grant FPU18/02 116).

In species that regenerate the injured spinal cord, the ependymal region is a source of new cells and a prominent coordinator of regeneration. In mammals, cells at the ependymal region proliferate in normal conditions and react after injury, but in humans, the central canal is lost in the majority of individuals from early childhood. It is replaced by a structure that does not proliferate after damage and is formed by large accumulations of ependymal cells, strong astrogliosis and perivascular pseudo-rosettes. We inform here of two additional mammals that lose the central canal during their lifetime: the Naked Mole-Rat (NMR, Heterocephalus glaber) and the mutant hyh (hydrocephalus with hop gait) mice. The morphological study of their spinal cords shows that the tissue substituting the central canal is not similar to that found in humans. In both NMR and hyh mice, the central canal is replaced by tissue reminiscent of normal lamina X and may include small groups of ependymal cells in the midline, partially resembling specific domains of the former canal. However, no features of the adult human ependymal remnant are found, suggesting that this structure is a specific human trait. In order to shed some more light on the mechanism of human central canal closure, we provide new data suggesting that canal patency is lost by delamination of the ependymal epithelium, in a process that includes apical polarity loss and the expression of signaling mediators involved in epithelial to mesenchymal transitions.

Background In obstructive congenital hydrocephalus, cerebrospinal fluid accumulation is associated with high intracranial pressure and the presence of periventricular edema, ischemia/hypoxia, damage of the white matter, and glial reactions in the neocortex. The viability and short time effects of a therapy based on bone marrow-derived mesenchymal stem cells (BM-MSC) have been evaluated in such pathological conditions in the hyh mouse model. Methods BM-MSC obtained from mice expressing fluorescent mRFP1 protein were injected into the lateral ventricle of hydrocephalic hyh mice at the moment they present a very severe form of the disease. The effect of transplantation in the neocortex was compared with hydrocephalic hyh mice injected with the vehicle and non-hydrocephalic littermates. Neural cell populations and the possibility of transdifferentiation were analyzed. The possibility of a tissue recovering was investigated using 1H High-Resolution Magic Angle Spinning Nuclear Magnetic Resonance (1H HR-MAS NMR) spectroscopy, thus allowing the detection of metabolites/osmolytes related with hydrocephalus severity and outcome in the neocortex. An in vitro assay to simulate the periventricular astrocyte reaction conditions was performed using BM-MSC under high TNFα level condition. The secretome in the culture medium was analyzed in this assay. Results Four days after transplantation, BM-MSC were found undifferentiated and scattered into the astrocyte reaction present in the damaged neocortex white matter. Tissue rejection to the integrated BM-MSC was not detected 4 days after transplantation. Hyh mice transplanted with BM-MSC showed a reduction in the apoptosis in the periventricular neocortex walls, suggesting a neuroprotector effect of the BM-MSC in these conditions. A decrease in the levels of metabolites/osmolytes in the neocortex, such as taurine and neuroexcytotoxic glutamate, also indicated a tissue recovering. Under high TNFα level condition in vitro, BM-MSC showed an upregulation of cytokine and protein secretion that may explain homing, immunomodulation, and vascular permeability, and therefore the tissue recovering. Conclusions BM-MSC treatment in severe congenital hydrocephalus is viable and leads to the recovery of the severe neurodegenerative conditions in the neocortex. NMR spectroscopy allows to follow-up the effects of stem cell therapy in hydrocephalus.

In congenital hydrocephalus, cerebrospinal fluid accumulation is associated with increased intracranial pressure (ICP), ischemia/hypoxia, metabolic impairment, neuronal damage, and astrocytic reaction. The aim of this study was to identify whether a metabolite profile revealing tissue responses according to the severity of hydrocephalus can be detected. The hyh mutant mouse used for this study exhibits 2 different forms of hydrocephalus, severe and moderate. In a comprehensive investigation into the 2 progressions of hydrocephalus, mice with severe hydrocephalus were found to have higher ICP and astrocytic reaction. Several metabolites from the mouse brain cortex were analyzed with 1H high-resolution magic angle spinning nuclear magnetic resonance (1H HR-MAS NMR) spectroscopy. A differential profile for metabolites including glutamate and glutamine was found to correlate with the severity of hydrocephalus and can be explained due to differential astrocytic reactions, neurodegenerative conditions, and the presence of ischemia. The glutamate transporter EAAT2 and the metabolite taurine were found to be key histopathological markers of affected parenchymata. In conclusion, a differential metabolite profile can be detected according to the severity of hydrocephalus and associated ICP and therefore can be used to monitor the efficacy of experimental therapies.

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A soluble N-ethylmaleimide-sensitive factor-attachment protein alpha (αSNAP) is a multifunctional scaffolding protein that regulates intracellular vesicle trafficking and signaling. In cultured intestinal epithelial cells, αSNAP has been shown to be essential for cell survival, motility, and adhesion; however, its physiologic functions in the intestinal mucosa remain unknown. In the present study, we used a mouse with a spontaneous hydrocephalus with hop gait (hyh) mutation of αSNAP to examine the roles of this trafficking protein in regulating intestinal epithelial homeostasis in vivo. Homozygous hyh mice demonstrated decreased expression of αSNAP protein in the intestinal epithelium, but did not display gross abnormalities of epithelial architecture in the colon and ileum. Such αSNAP depletion attenuated differentiation of small intestinal epithelial enteroids ex vivo. Furthermore, αSNAP-deficient mutant animals displayed reduced formation of lysozyme granules in small intestinal crypts and decreased expression of lysozyme and defensins in the intestinal mucosa, which is indicative of defects in Paneth cell differentiation. By contrast, development of Goblet cells, enteroendocrine cells, and assembly of enterocyte apical junctions was not altered in hyh mutant mice. Our data revealed a novel role of αSNAP in the intestinal Paneth cell differentiation in vivo.

et al., Lysophosphatidic acid (LPA) is an intercellular signaling lipid that regulates multiple cellular functions, acting through specific G-protein coupled receptors (LPA1–6). Our previous studies using viable Malaga variant maLPA1-null mice demonstrated the requirement of the LPA1 receptor for normal proliferation, differentiation, and survival of the neuronal precursors. In the cerebral cortex LPA1 is expressed extensively in differentiating oligodendrocytes, in parallel with myelination. Although exogenous LPA-induced effects have been investigated in myelinating cells, the in vivo contribution of LPA1 to normal myelination remains to be demonstrated. This study identified a relevant in vivo role for LPA1 as a regulator of cortical myelination. Immunochemical analysis in adult maLPA1-null mice demonstrated a reduction in the steady-state levels of the myelin proteins MBP, PLP/DM20, and CNPase in the cerebral cortex. The myelin defects were confirmed using magnetic resonance spectroscopy and electron microscopy. Stereological analysis limited the defects to adult differentiating oligodendrocytes, without variation in the NG2+ precursor cells. Finally, a possible mechanism involving oligodendrocyte survival was demonstrated by the impaired intracellular transport of the PLP/DM20 myelin protein which was accompanied by cellular loss, suggesting stress-induced apoptosis. These findings describe a previously uncharacterized in vivo functional role for LPA1 in the regulation of oligodendrocyte differentiation and myelination in the CNS, underlining the importance of the maLPA1-null mouse as a model for the study of demyelinating diseases., This work was supported by the Carlos III Health Institute, State Department of Research, Development and Innovation, Spanish Ministry of Economy and Competitiveness (Grant Numbers PI10/02514—co-funded by European Research Development Fund—, to G.E-T.; SAF2011 to IV-N); Andalusian Regional Ministries of Health (Nicolás Monardes Programme, and Grants PI0187/2008, PI0232/2007 to G.E-T.) and of Economy, Innovation, Science and Employment (CTS643 and CTS433 research group grants to G.E-T. and F.R-DF., respectively); Ramon Areces Foundation (Ramon Areces Fellowship to B.G-D.); and the National Institutes of Health (USA) (Grant Numbers MH051699 and MH01723 to J.C.).

espanolIntroduccion El dolor lumbar cronico tiene una alta prevalencia en los paises industrializados, donde es una de las principales causas de incapacidad laboral. Con frecuencia, los pacientes con dolor lumbar cronico en tratamiento con opiaceos sufren episodios de dolor irruptivo, pero los datos de prevalencia y preferencias de tratamiento son escasos. La prevalencia, caracteristicas y manejo del dolor irruptivo de pacientes con dolor cronico de origen lumbar en Andalucia y Melilla es desconocida. Objetivos 1. Evaluacion de la prevalencia del dolor irruptivo en pacientes con dolor cronico secundario a lumbalgia cronica en Andalucia y Melilla (objetivo primario). 2. Caracterizacion del dolor irruptivo en pacientes con dolor cronico secundario a lumbalgia cronica basado en la etiologia, patologia, asi como en otras caracteristicas clinicas. 3. Evaluacion de la prevalencia de cada una de las diferentes causas de dolor. 4. Identificar las posibles asociaciones entre los diferentes tipos de dolor irruptivo y aspectos sociodemograficos, clinicos y asistenciales. Material y metodos En el estudio de prevalencia del dolor irruptivo participaron 1.868 pacientes y en el de caracterizacion 295 pacientes. En el estudio de prevalencia se recogieron las siguientes variables: presencia de dolor irruptivo, sexo y edad. En el estudio de caracterizacion se recogieron datos relativos al tipo y localizacion del dolor, tratamiento, cumplimiento y satisfaccion del paciente. Resultados La prevalencia del dolor irruptivo en pacientes con dolor cronico secundario a lumbalgia cronica es del 37,5 % (IC 95 %: 35,3-39,7 %), y es similar en hombres y mujeres. Un 75 % de los pacientes son mayores de 50 anos. La media de dolor irruptivo fue de 84,4 puntos en la escala visual analogica (EVA). El tratamiento del dolor cronico es tratado con una amplia gama de opiaceos. El farmaco preferente de los pacientes que controlan el dolor irruptivo con opiaceos es fentanilo (78,3 %) y la forma de administracion mas comun de este es la nasal (53,2 %). El grado de cumplimiento es alto y un 46,3 % de los pacientes consideran muy satisfactorio el control de su dolor irruptivo. Conclusiones Los datos epidemiologicos sobre el dolor irruptivo en Andalucia y Melilla generados por este estudio nos ha permitido conocer su prevalencia y caracteristicas, asi como los tratamientos preferidos y el grado de satisfaccion de los pacientes. EnglishIntroduction Chronic low back pain is highly prevalent in industrialized countries, where it is one of the main causes of incapacity for work. Patients with chronic low back pain in treatment with opioids often experience episodes of breakthrough pain, but prevalence and treatment preferences data are scarce. The prevalence, characteristics, and management of breakthrough pain of patients with chronic pain of lumbar origin in Andalusia and Melilla are unknown. Objectives 1. Evaluation of the prevalence of breakthrough pain in patients with chronic pain secondary to chronic low back pain in Andalusia and Melilla (primary endpoint). 2. Characterization of breakthrough pain in patients with chronic pain secondary to chronic low back pain based on etiology, pathology, and other clinical characteristics. 3. Assessment of the prevalence of each of the different causes of pain. 4. Identify the possible associations between different types of breakthrough pain and sociodemographic, clinical and healthcare factors. Materials and methods 1,868 patients participated in the study of prevalence of breakthrough pain and 295 patients participated in the characterization study. In the prevalence study the following variables were collected: presence of breakthrough pain, sex and age. In the characterization study data were collected regarding the type and location of pain, treatment, compliance, and patient satisfaction. Results: The prevalence of breakthrough pain in patients with chronic pain secondary to chronic low back pain is 37.5% (95 % CI: 35.3-39.7%), and is similar in men and women. 75% of the patients are older than 50 years. The mean value of breakthrough pain was 84.4 points in a visual analogue scale (VAS). Chronic lumbar pain is treated with a wide range of opiates. The preferred drug of patients who control breakthrough pain with opioids is fentanyl (78.3%) and its most common form of administration is nasal (53.2%). The degree of compliance is high and 46.3 % of patients consider the control of their breakthrough pain very satisfactory. Conclusions: Epidemiological data on the breakthrough pain in Andalusia and Melilla generated by this study has allowed us to know its prevalence and characteristics, as well as the preferred treatments and the degree of satisfaction of the patients.

Most cells of the developing mammalian brain derive from the ventricular (VZ) and the subventricular (SVZ) zones. The VZ is formed by the multipotent radial glia/neural stem cells (NSCs) while the SVZ harbors the rapidly proliferative neural precursor cells (NPCs). Evidence from human and animal models indicates that the common history of hydrocephalus and brain maldevelopment starts early in embryonic life with disruption of the VZ and SVZ. We propose that a "cell junction pathology" involving adherent and gap junctions is a final common outcome of a wide range of gene mutations resulting in proteins abnormally expressed by the VZ cells undergoing disruption. Disruption of the VZ during fetal development implies the loss of NSCs whereas VZ disruption during the perinatal period implies the loss of ependyma. The process of disruption occurs in specific regions of the ventricular system and at specific stages of brain development. This explains why only certain brain structures have an abnormal development, which in turn results in a specific neurological impairment of the newborn. Disruption of the VZ of the Sylvian aqueduct (SA) leads to aqueductal stenosis and hydrocephalus, while disruption of the VZ of telencephalon impairs neurogenesis. We are currently investigating whether grafting of NSCs/neurospheres from normal rats into the CSF of hydrocephalic mutants helps to diminish/repair the outcomes of VZ disruption.

[EN] Virtual laboratories are a very powerful tool to help in transmitting scientific knowledge. Within the actual framework of university teaching, such tools are of great value due to its capabilities of self-teaching to the students of technical studies. In the present work, the authors describe a virtual laboratory developed with the Graphical User Interface (Gui) of the MatlabQc package. The aim of this virtual laboratory is to obtain the diagrams (i.e., graphical representation) of axial forces, shear forces and bending moments in plane, two-dimensional frames., [ES] Los laboratorios virtuales constituyen una herramienta de gran potencial para la transmisión del conocimiento científico. En el marco de las enseñanzas universitarias actuales, dichas herramientas adquieren una importancia capital dadas las posibilidades de autoaprendizaje que ofrecen a los estudiantes de los títulos técnicos de grado. En este marco, el presente trabajo describe un laboratorio virtual desarrollado con la interfaz gráfica de usuario (Gui) de MatlabQc , que permite obtener los diagramas de esfuerzos axiles, cortantes y flectores en estructuras aporticadas isostáticas planas, con las cargas contenidas en su propio plano.

α-SNAP is an essential component of the protein machinery responsible for membrane fusion events in different cell types. The hyh (hydrocephalus with hop gait) mouse carries a missense mutation in Napa gene that results in a point mutation (M105I) in α-SNAP protein. Homozygous animals for the mutant allele have been identified by the clinical and/or neuropathological phenotype, or by direct sequencing of PCR products. The aims of the present study were (i) to develop a high-throughput technique to genotype hyh mice, (ii) to correlate genotype-phenotype, and (iii) to analyze the earliest pathological changes of hyh mutant mice. As no restriction sites are affected by the hyh mutation, we resolved this problem by creating a BspHI restriction site with a modified (mismatch) polymerase chain reaction (PCR) primer in wild-type allele. This artificially created restriction site (ACRS)–PCR technique is a simple, rapid and reliable method to genotype hyh mice in a day-work procedure. Biochemical and histological analysis of genotyped hyh embryos at different developmental stages allowed us to identify and characterize the earliest brain pathological changes of the hyh phenotype, including the first signs of neuroepithelial disruption and neuronal ectopia. In addition, genotype-phenotype analysis of 327 animals confirmed that (i) hyh is a single-gene autosomal recessive disorder, and (ii) the disorder has 100% penetrance (i.e., the mutation was only present in affected mice). The genotyping method described here enhances the potentiality of hyh mouse as a unique in vivo model to study the role of membrane trafficking in different developmental and physiological processes.

The endophytic phase of Pseudomonas savastanoi pv. savastanoi in olive stems and the structural and ultrastructural histogenesis of olive knots have been studied. Construction of a stable plasmid vector expressing the green fluorescent protein, in combination with the use of in vitro olive plants, allowed real-time monitoring of P. savastanoi pv. savastanoi infection. The infection process was also examined by bright field and epifluorescence microscopy as well as by scanning and transmission electron microscopy. Hypertrophy of the stem tissue was concomitant with the formation of bacterial aggregates, microcolonies and multilayer biofilms, over the cell surfaces and the interior of plasmolysed cells facing the air-tissue interface of internal opened fissures, and was followed by invasion of the outer layers of the hypertrophied tissue. Pathogenic invasion of the internal lumen of newly formed xylem vessels, which were connected with the stem vascular system, was also observed in late stages of infection. Ultrastructural analysis of knot sections showed the release of outer membrane vesicles from the pathogen surface, a phenomenon not described before for bacterial phytopathogens during host infection. This is the first real-time monitoring of P. savastanoi disease development and the first illustrated description of the ultrastructure of P. savastanoi-induced knots.

To determine if ventricular zone (VZ) and subventricular zone (SVZ) alterations are associated with intraventricular hemorrhage (IVH) and posthemorrhagic hydrocephalus, we compared postmortem frontal and subcortical brain samples from 12 infants with IVH and 3 nonneurological disease controls without hemorrhages or ventriculomegaly. Birth and expiration estimated gestational ages were 23.0-39.1 and 23.7-44.1 weeks, respectively; survival ranges were 0-42 days (median, 2.0 days). Routine histology and immunohistochemistry for neural stem cells (NSCs), neural progenitors (NPs), multiciliated ependymal cells (ECs), astrocytes (AS), and cell adhesion molecules were performed. Controls exhibited monociliated NSCs and multiciliated ECs lining the ventricles, abundant NPs in the SVZ, and medial vs. lateral wall differences with a complex mosaic organization in the latter. In IVH cases, normal VZ/SVZ areas were mixed with foci of NSC and EC loss, eruption of cells into the ventricle, cytoplasmic transposition of N-cadherin, subependymal rosettes, and periventricular heterotopia. Mature AS populated areas believed to be sites of VZ disruption. The cytopathology and extension of the VZ disruption correlated with developmental age but not with brain hemorrhage grade or location. These results corroborate similar findings in congenital hydrocephalus in animals and humans and indicate that VZ disruption occurs consistently in premature neonates with IVH.

Under normal physiological conditions the cerebrospinal fluid (CSF) is secreted continuously, although this secretion undergoes circadian variations. Mechanisms operating at the vascular side of the choroidal cells involve a sympathetic and a cholinergic innervation, with the former inhibiting and the latter stimulating CSF secretion. There are also regulatory mechanisms operating at the ventricular side of the choroidal cells, where receptors for monoamines such as dopamine, serotonin, and melatonin, and for neuropeptides such as vasopressin, atrial natriuretic hormone, and angiotensin II, have been identified. These compounds, that are normally present in the CSF, participate in the regulation of CSF secretion. Although the mechanisms responsible for the CSF circulation are not fully understood, several factors are known to play a role. There is evidence that the subcommissural organ (SCO)–Reissner's fiber (RF) complex is one of the factors involved in the CSF circulation. In mammals, the predominant route of escape of CSF into blood is through the arachnoid villi. In lower vertebrates, the dilatation of the distal end of the central canal, known as terminal ventricle or ampulla caudalis, represents the main site of CSF escape into blood. Both the function and the ultrastructural arrangement of the ampulla caudalis suggest that it may be the ancestor structure of the mammalian arachnoid villi. RF-glycoproteins reaching the ampulla caudalis might play a role in the formation and maintenance of the route communicating the CSF and blood compartments. The SCO-RF complex may participate, under physiological conditions, in the circulation and reabsorption of CSF. Under pathological conditions, the SCO appears to be involved in the pathogeneses of congenital hydrocephalus. Changes in the SCO have been described in all species developing congenital hydrocephalus. In these reports, the important question whether the changes occurring in the SCO precede hydrocephalus, or are a consequence of the hydrocephalic state, has not been clarified. Recently, evidence has been obtained indicating that a primary defect of the SCO-RF complex may lead to hydrocephalus. Thus, a primary and selective immunoneutralization of the SCO-RF complex during the fetal and early postnatal life leads to absence of RF, aqueductal stenosis, increased CSF concentration of monoamines, and a moderate but sustained hydrocephalus. Microsc. Res. Tech. 52:591–607, 2001. © 2001 Wiley-Liss, Inc.

Stenosis of the cerebral aqueduct seems to be a key event for the development of congenital hydrocephalus. The causes of such a stenosis are not well known. Overholser et al. in 1954 (Anat Rec 120:917-933) proposed the hypothesis that a dysfunction of the subcommissural organ (SCO) leads to aqueductal stenosis and congenital hydrocephalus. The SCO is a brain gland, located at the entrance of the cerebral aqueduct, that secretes glycoproteins into the cerebrospinal fluid that, upon release, assemble into a fibrous structure known as Reissner's fiber (RF). By the permanent addition of new molecules to its rostral end, RF grows and extends along the aqueduct, fourth ventricle, and central canal of the spinal cord. The immunological blockage of the SCO-RF complex has been used to test Overholser's hypothesis. The following was the sequence of events occurring in pregnant rats that had been immunized with RF glycoproteins: the mother produced anti-RF antibodies and transferred them to the fetus through the placenta and to the pup through the milk, and the antibodies reached the brain of the fetus and pup and blocked the SCO-RF complex. This resulted in a permanent absence of RF that was followed by stenosis of the cerebral aqueduct, and then by the appearance of hydrocephalus. The latter was patent until the end of the 6-month observation period. The chronic hydrocephalic state appeared, in turn, to induce new alterations of the SCO. It is concluded that a selective immunological knock out of the SCO-RF complex leads to hydrocephalus.

The neuroepithelium is a germinal epithelium containing progenitor cells that produce almost all of the central nervous system cells, including the ependyma. The neuroepithelium and ependyma constitute barriers containing polarized cells covering the embryonic or mature brain ventricles, respectively; therefore, they separate the cerebrospinal fluid that fills cavities from the developing or mature brain parenchyma. As barriers, the neuroepithelium and ependyma play key roles in the central nervous system development processes and physiology. These roles depend on mechanisms related to cell polarity, sensory primary cilia, motile cilia, tight junctions, adherens junctions and gap junctions, machinery for endocytosis and molecule secretion, and water channels. Here, the role of both barriers related to the development of diseases, such as neural tube defects, ciliary dyskinesia, and hydrocephalus, is reviewed.

TETRATRICOPEPTIDE THIOREDOXIN-LIKE (TTL) proteins are characterized by the presence of six tetratricopeptide repeats in conserved positions and a carboxyl-terminal region known as the thioredoxin-like domain with homology to thioredoxins. In Arabidopsis (Arabidopsis thaliana), the TTL gene family is composed by four members, and the founder member, TTL1, is required for osmotic stress tolerance. Analysis of sequenced genomes indicates that TTL genes are specific to land plants. In this study, we report the expression profiles of Arabidopsis TTL genes using data mining and promoter-reporter β-glucuronidase fusions. Our results show that TTL1, TTL3, and TTL4 display ubiquitous expression in normal growing conditions but differential expression patterns in response to osmotic and NaCl stresses. TTL2 shows a very different expression pattern, being specific to pollen grains. Consistent with the expression data, ttl1, ttl3, and ttl4 mutants show reduced root growth under osmotic stress, and the analysis of double and triple mutants indicates that TTL1, TTL3, and TTL4 have partially overlapping yet specific functions in abiotic stress tolerance while TTL2 is involved in male gametophytic transmission.

Dystrophic neurites associated with amyloid plaques precede neuronal death and manifest early in Alzheimer's disease (AD). In this work we have characterized the plaque-associated neuritic pathology in the hippocampus of young (4- to 6-month-old) PS1(M146L)/APP(751SL) mice model, as the initial degenerative process underlying functional disturbance prior to neuronal loss. Neuritic plaques accounted for almost all fibrillar deposits and an axonal origin of the dystrophies was demonstrated. The early induction of autophagy pathology was evidenced by increased protein levels of the autophagosome marker LC3 that was localized in the axonal dystrophies, and by electron microscopic identification of numerous autophagic vesicles filling and causing the axonal swellings. Early neuritic cytoskeletal defects determined by the presence of phosphorylated tau (AT8-positive) and actin-cofilin rods along with decreased levels of kinesin-1 and dynein motor proteins could be responsible for this extensive vesicle accumulation within dystrophic neurites. Although microsomal A beta oligomers were identified, the presence of A11-immunopositive A beta plaques also suggested a direct role of plaque-associated A beta oligomers in defective axonal transport and disease progression. Most importantly, presynaptic terminals morphologically disrupted by abnormal autophagic vesicle buildup were identified ultrastructurally and further supported by synaptosome isolation. Finally, these early abnormalities in axonal and presynaptic structures might represent the morphological substrate of hippocampal dysfunction preceding synaptic and neuronal loss and could significantly contribute to AD pathology in the preclinical stages.

[EN] In this work, we present a graphic user interface (Gui) developed in MatlabQc to easily obtain the gravity center and the matrix of inertia of a plane domain. It is allowed to include holes, an also to consider curved or straight edges. The application represents graphically the domain, the position of the centre of gravity and the principal axes of inertia. From a pedagogical point of view, it can be usefully employed in the context of different subjects in university teaching, such as: Physics, Strength of Materials, Structural Analysis, etc., [ES] En este trabajo presentamos una interfaz gráfica de usuario de MatlabQc que permite obtener fácilmente el centro de gravedad y la matriz de inercia de un dominio plano, con posibilidad de agujeros y cuyo borde esté formado por líneas poligonales y arcos de circunferencia. Además la aplicación representa gráficamente el dominio, la posición del centro de gravedad y los ejes principales de inercia. Desde un punto de vista pedagógico puede ser útil en diversas asignaturas de carreras de ingeniería como física, resistencia de materiales, cálculo de estructuras, etc.

Summary The endophytic phase of Pseudomonas savastanoi pv. savastanoi in olive stems and the structural and ultrastructural histogenesis of olive knots have been studied. Construction of a stable plasmid vector expressing the green fluorescent protein, in combination with the use of in vitro olive plants, allowed real‐time monitoring of P. savastanoi pv. savastanoi infection. The infection process was also examined by bright field and epifluorescence microscopy as well as by scanning and transmission electron microscopy. Hypertrophy of the stem tissue was concomitant with the formation of bacterial aggregates, microcolonies and multilayer biofilms, over the cell surfaces and the interior of plasmolysed cells facing the air‐tissue interface of internal opened fissures, and was followed by invasion of the outer layers of the hypertrophied tissue. Pathogenic invasion of the internal lumen of newly formed xylem vessels, which were connected with the stem vascular system, was also observed in late stages of infection. Ultrastructural analysis of knot sections showed the release of outer membrane vesicles from the pathogen surface, a phenomenon not described before for bacterial phytopathogens during host infection. This is the first real‐time monitoring of P. savastanoi disease development and the first illustrated description of the ultrastructure of P. savastanoi‐induced knots.

[EN] Computer-assisted Finite Element Method is a widespread tool in the field of engineering. In certain applications, it is necessary to modify the mesh of the model during the analysis, which is known as ’remeshing’. In these situations, it is usually needed to perform some measurements, p.e., to calculate the volume of a region. Commercial packages can cover these needed, but they reveal to be limited in some situations. In this work, an application developed in MatlabQc is presented to approximately calculate the volume of a spatial region comprised between to meshes. The first is a quadrilateral finite element mesh, and the second mesh is topologically equivalent to the first and obtained through a remeshing process., [ES] El Método de los Elementos Finitos asistido por ordenador es una herramienta muy extendida en ingeniería. En ciertas aplicaciones, se precisa modificar la malla del modelo a lo largo del proceso de simulación, lo que se conoce como remallado. En estos casos, además, suele ser necesario realizar mediciones en el modelo de elementos finitos, como por ejemplo calcular el volumen de ciertas regiones del mismo. Los paquetes comerciales disponibles en la actualidad permiten satisfacer algunas de estas necesidades, aunque adolecen de ciertas limitaciones. En este trabajo se presenta una aplicación desarrollada en el código propio de MatlabQc, que permite el cálculo aproximado del volumen de una región del espacio comprendida entre una malla de elementos finitos cuadriláteros lineales y otra malla topológicamente equivalente a la anterior, esta última obtenida a través de un proceso de remallado.

A heterogeneous population of ependymal cells lines the brain ventricles. The evidence about the origin and birth dates of these cell populations is scarce. Furthermore, the possibility that mature ependymal cells are born (ependymogenesis) or self-renewed (ependymal proliferation) postnatally is controversial. The present study was designed to investigate both phenomena in wild-type (wt) and hydrocephalic α-SNAP mutant (hyh) mice at different postnatal stages. In wt mice, proliferating cells in the ventricular zone (VZ) were only found in two distinct regions: the dorsal walls of the third ventricle and Sylvian aqueduct (SA). Most proliferating cells were monociliated and nestin+, likely corresponding to radial glial cells. Postnatal cumulative BrdU-labeling showed that most daughter cells remained in the VZ of both regions and they lost nestin-immunoreactivity. Furthermore, some labeled cells became multiciliated and GLUT-1+, indicating they were ependymal cells born postnatally. Postnatal pulse BrdU-labeling and Ki-67 immunostaining further demonstrated the presence of cycling multiciliated ependymal cells. In hydrocephalic mutants, the dorsal walls of the third ventricle and SA expanded enormously and showed neither ependymal disruption nor ventriculostomies. This phenomenon was sustained by an increased ependymogenesis. Consequently, in addition to the physical and geometrical mechanisms traditionally explaining ventricular enlargement in fetal-onset hydrocephalus, we propose that postnatal ependymogenesis could also play a role. Furthermore, as generation of new ependymal cells during postnatal stages was observed in distinct regions of the ventricular walls, such as the roof of the third ventricle, it may be a key mechanism involved in the development of human type 1 interhemispheric cysts.

Background In human foetal hydrocephalus and spina bifida aperta (SBA), the pathogenesis of abnormal cortical development is poorly understood. Previous reports have shown that neuroepithelial/ependymal denudation is involved in the neuropathogenesis of human foetal hydrocephalus and SBA. Interestingly, loss of the neuroepithelium/ ependyma (denudation) at the Sylvian aqueduct is preceded by defective expression of adherent and gap junction proteins. In human foetal hydrocephalus, we aimed to investigate whether abnormal cortical development is similarly associated with intercellular ependymal defects at the telencephalic (sub)ventricular zones.

Pseudomonas savastanoi pv. savastanoi strain NCPPB 3335 is a model bacterial pathogen for studying the molecular basis of disease production in woody hosts. We report the sequencing of the hrpS-to-hrpZ region of NCPPB 3335, which has allowed us to determine the phylogenetic position of this pathogen with respect to previously sequenced Pseudomonas syringae hrp clusters. In addition, we constructed a mutant of NCPPB 3335, termed T3, which carries a deletion from the 3′ end of the hrpS gene to the 5′ end of the hrpZ operon. Despite its inability to multiply in olive tissues and to induce tumor formation in woody olive plants, P. savastanoi pv. savastanoi T3 can Induce knot formation on young micropropagated olive plants. However, the necrosis and formation of internal open cavities previously reported in knots induced by the wild-type strain were not observed in those induced by P. savastanoi pv. savastanoi T3. Tagging of P. savastanoi pv. savastanoi T3 with green fluorescent protein (GFP) allowed real-time monitoring of its behavior on olive plants. In olive plant tissues, the wild-type strain formed aggregates that colonized the intercellular spaces and internal cavities of the hypertrophic knots, while the mutant T3 strain showed a disorganized distribution within the parenchyma of the knot. Ultrastructural analysis of knot sections revealed the release of extensive outer membrane vesicles from the bacterial cell surface of the P. savastanoi pv. savastanoi T3 mutant, while the wild-type strain exhibited very few vesicles. This phenomenon has not been described before for any other bacterial phytopathogen during host infection. Copyright © 2010, American Society for Microbiology. All Rights Reserved., This research has been supported by Spanish Plan Nacional I D I grants AGL2008-55311-CO2-01 and AGL2008-55311-CO2-02, cofi- nanced by FEDER, as well as by Integrated Action Spain-Italy HI01-81 and by Ente Cassa di Risparmio di Firenze (2007.1005 and 2008.1573). I.P.-M. was awarded a Spanish MEC fellowship (AP2002- 3800), and L.L. and I.M.M. were supported by Carlsberg Foundation (Denmark) grant ANS-1282/20 and the Ramo ́n Areces Foundation (Spain), respectively.

Neural stem cells persist after embryonic development in the subventricular zone (SVZ) niche and produce new neural cells during postnatal life; ependymal cells are a key component associated with this neurogenic niche. In the animal model of human hydrocephalus, the hyh mouse, the ependyma of the lateral ventricles is progressively lost during late embryonic and early postnatal life and disappears from most of the ventricular surface throughout its life span. To determine the potential consequences of this loss on the SVZ, we characterized the abnormalities in this neurogenic niche in hyh mice. There was overall disorganization and a marked reduction of proliferative cells in the SVZ of both newborn and adult hyh hydrocephalic mice in vivo; neuroblasts were displaced to the ventricular surface, and their migration through the rostral migratory stream was reduced. The numbers of resident neural progenitor cells in hyh mice were also markedly reduced, but they were capable of proliferating, forming neurospheres, and differentiating into neurons and glia in vitro in a manner indistinguishable from that of wild-type progenitor cells. These findings suggest that the reduction of proliferative activity observed in vivo is not caused by a cell autonomous defect of SVZ progenitors but is a consequence of a reduced number of these cells. Furthermore, the overall tissue disorganization of the SVZ and displacement of neuroblasts imply alterations in the neurogenic niche of postnatal hyh mice.

BACKGROUND: Despite the excellent prognostic of chronic myeloid leukemia (CML) patients since the introduction of tyrosine kinase inhibitors (TKIs), approximately 50% of patients that are treated with TKIs will discontinue first line treatment due to lack of efficacy or intolerance. Once patients need a second line treatment, a considerable proportion of patients will need third or even fourth line therapy during further evolution. At this moment, there is a lack of data about real benefit of this group of patients. We have recently published our experience of 30 CML patients treated with bosutinib in 4th line. We present an update of the study where we have increased the number of patients, and the follow-up. The aim of this study is to present safety and efficacy data CML chronic phase patients treated with bosutinib in 4th line. METHODS: We have collected data from 59 CML patients treated with bosutinib in 4th line after resistance or intolerance to IM, NI and DA. 51 patients have been treated under the Spanish compassionate use program (36 centers) and 10 patients were treated in a single institution from United Kingdom. Median age of patients at diagnosis was 53 years. The percentage of low, intermediate and high risk Sokal groups were 47%, 37% and 16%. Median time TKIs exposure before bosutinib was 9 years. The most common indication (30/59) was intolerant to DA and NI. Patients' dispositions and main line characteristics are shown in table 1. RESULTS: Median follow-up was 14.3 months. All patients started bosutinib at 500mg/d, median dose of was 450mg/d. Overall probabilities to either achieve or maintain previous response were 96% (57/59), 62% (37/59), 40% (24/59) and 17% (10/59) for complete hematological response (CHR), complete cytogenetic response (CCyR), major molecular response (MMR) and MR4.5 respectively. However, probabilities to obtain responses (in patients without response evaluated at baseline) were 27% (7/26), 26% (12/45) and 12% (7/55) for CCyR, MMR and MR4.5. As expected, probabilities to obtain CCyR were lower for patients resistant to DA and NI patients than for patients intolerant to DA and NI (8% VS 44%). Event free survival (EFS) and progression free survival (PFS) probabilities were 50% and 83% by 27 month. Treatment was discontinued in 20/58 (34%), most frequent reasons being adverse events 9/59(15%), lack of efficacy 5/59 (8.5%), disease progression 2/59 (3.4%) and death 1/59 (1.7%). Two patients discontinued due to stem cell transplantation. The adverse events that led to treatment discontinuation were pleural effusion (3), diarrhea (2), rash, renal impairment, auricular fibrillation and liver enzyme elevation one patient each. Overall, bosutinib was well tolerated. Grade 3-4 hematological toxicities were 3%, 6% and 6% for anemia, thromboctytopenia and neutropenia. Most common non hematological side effects were diarrhea (39%, nauseas 13% and liver alterations 14% and pleural effusion 14%. CONCLUSIONS: Little is known about the therapeutic role of Bosutinib in 4th line. The series presented here is, to our knowledge, the largest being presented. Bosutinib seems to be an appropriate treatment option for patients resistant or intolerant to three prior TKIs. | | IM+NI-I+DA-R (N=4) | IM+NI-R+DA-R (N=18) | IM+NI-I+DA-I (N=30) | IM+NI-R+DA-I (N=7) | Total (N=59) | | ------------------------------------------------ | ----------------------- | ------------------------ | ----------------------- | ------------------------ | ------------------------- | ------------------------- | | Sex, N (%) Male | 2 (50) | 11 (61.1) | 16 (53.3) | 2 (28.6) | 31 (52.5) | | Median age of diagnosis, yr (range) | 57.32 (50-64) | 49.19 (23-73) | 54.95 (21-89) | 48.87 (26-68) | 53.15 (21-89) | | Median age of Bosutinib initiation, yr (range) | 69.13 (61-70) | 62.27 (39-79) | 64.85 (25-90) | 64.79(35-74) | 63.7 (25-9) | | Median follow up, months (range) | 18.5(7.8-34.1) | 8.4(1.22-36.1) | 16.3(0.5-34.7) | 23.4(3.3-28.9) | 14.3(0.7-36.1) | | SOKAL Index at diagnosis, N (%) | High | 2(50.0) | 4 (23.5) | 1 (4.3) | 1 (20) | 8 (16.3) | | Intermediate | 1 (25.0) | 5 (29.4) | 10(43.5) | 2 (40) | 18 (36.7) | | Low | 1 (25.0) | 8 (47.1) | 12 (52.2) | 2 (40) | 23 (46.9) | | Median Time from first TKI to BOS, (yr, range) | 10.3 (4.8-11.9) | 9.3 (2.0-11.4) | 8.8 (0.7-13.6) | 8.2 (5.1-12.3) | 8.8 (0.7-13.6) | | Median duration of prior therapy, months (range) | Imatinib | 38.8 (11.8-69.8) | 32.6 (6.3-96.8) | 26.2 (1.6-102.6) | 23.1 (8.3-66.8) | 28.8 (1.6-102.6) | | Dasatinib | 21.5 (12.6-75) | 21.8 (7.7-69) | 31.4 (0.4-87.1) | 23.7 (10.3-53.6) | 23.44 (0.4-87.1) | | Nilotinib | 19.1 (2.1-46.2) | 16.7 (5-65.6) | 8.9 (0.2-58.5) | 30.9 (6.9-49.3) | 14.3 (0.2-65.6) | * BOS: bosutinib, IM, imatinib; DA, dasatinib; NI, nilotinib, I: Intolerance, R: Resistant, Yr: year Table 1. Disclosures Garcia-Gutierrez: Ariad: Consultancy; Pfizer: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Novartis: Consultancy, Honoraria. Milojkovic: Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Ariad: Consultancy, Honoraria; BMS: Consultancy, Honoraria. Boque: Novartis: Honoraria; BMS: Honoraria; Celgene: Honoraria. Casado: Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Roche: Honoraria, Research Funding. Jimenez: Pfizer: Consultancy, Honoraria. Giraldo: Pfizer: Consultancy. Steegmann: Novartis: Consultancy, Honoraria, Research Funding, Speakers Bureau; BMS: Consultancy, Honoraria, Research Funding, Speakers Bureau; Pfizer: Consultancy, Honoraria, Research Funding, Speakers Bureau; Ariad: Consultancy, Honoraria, Research Funding, Speakers Bureau.

Background A population (30%) of hydrocephalic hyh mutant mice develop a slowly progressive hydrocephalus and survive for periods ranging between 2 months and 2 years. Certain characteristics of these mice, such as time of onset of hydrocephalus, type of abnormality of CSF dynamics, clinical evolution and survival/death rate, resemble several types of human congenital hydrocephalus. They represent an exceptional animal model to investigate neuropathological and physiopathological aspects of a brain "adapting" to a virtually life-lasting hydrocephalus

The hyh mouse carrying a point mutation in the gene encoding for soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein alpha (alpha-SNAP) develops inherited hydrocephalus. The investigation was designed to study: (i) the clinical evolution of hyh mice; (ii) factors other than the alpha-SNAP mutation that may influence the expression of hydrocephalus; (iii) the neuropathological features underlying the different forms of clinical evolution. The study included 3017 mice, 22.4% of which were hydrocephalic. The neuropathological study was performed in 112 mice by use of light and electron microscopy. It was found that maternal- and sex-related factors are involved in the heterogeneous expression of hyh phenotype. The clinical evolution recorded throughout a 4-year period also revealed a heterogeneous expression of the hydrocephalic phenotype. Two subpopulations were distinguished: (i) 70% of mice underwent a rapidly progressive hydrocephalus and died during the first 2 months of life; they presented macrocephaly, extremely large expansion of the ventricles, equilibrium impairment and decreased motor activity. (ii) Mice with slowly progressive hydrocephalus (30%) survived for periods ranging between 2 months and 2 years. They had no or moderate macrocephaly; moderate ventricular dilatation and preserved general motor activity; they all presented spontaneous ventriculostomies communicating the ventricles with the subarachnoid space, indicating that such communications play a key role in the long survival of these mice. The hyh mutant represents an ideal animal model to investigate how do the brain "adapt" to a virtually life-lasting hydrocephalus.

The hyh (hydrocephalus with hop gait) mutant micedevelop inherited hydrocephalus. A key feature in thismutant is that there is a foetal-onset ependymal denuda-tion which precedes cerebral aqueduct obliteration andhydrocephalus [1]. Recently, a point mutation in alpha-SNAP protein has been identified as responsible of thehyh phenotype [2]. However, preliminary findings fromour laboratory have suggested clinical and pathologicalheterogeneity in the expression of hydrocephalus, indicat-ing that other (nongenetic?) factors may influence thedegree of severity of this pathology. This is in accordancewith findings in other hydrocephalic mutant strains [3,4].The present investigation was designed to (a) study theclinical evolution of hydrocephalic mice in order to eval-uate wether or not clinical heterogeneity does actuallyoccur, (b) identify nongenetic factors (maternal age, mul-tiparity) that may affect such an evolution, and (c) iden-tify neuropathologic events underlying clinicalheterogeneity.

mutant mice suffer a congenital hydrocephalus trig-gered by ependyma denudation [1]. The ventricular sur-face in non hydrocephalic newborn mice is lined by theimmature ependyma, which is characterized for beingvimentin (-) and S100β (-), at variance in the adult ani-mals the mature ependyma expresses vimentin and S100 β[2]. On the other hand, in the hydrocephalic mice theependyma begins to denudate on the 12th day of gesta-tion, and at PN8 only some areas of lateral ventricle arestill endowed with ependyma. In parallel, astroglia startsto cover the denuded surface forming a new cell layer, theglial scar, which lines the damaged ventricular surface. Wehave studied the permeability to horseradish peroxidase(HRP) of these four regions at the ventricular wills:mature ependyma, and denuded areas with or withoutglial scar.

Dopamine receptors have been found in certain populations of non-neuronal cells in the brain, viz., discrete areas of ciliated ependyma and the ependymal cells of the choroid plexus. We have studied the presence of both tyrosine-hydroxylase-immunoreactive nerve fibers and dopamine receptors in the subcommissural organ (SCO), an ependymal brain gland that is located in the roof of the third ventricle and that secretes, into the cerebrospinal fluid, glycoproteins that aggregate to form Reissner’s fiber (RF). Antibodies against D2, D3, D4, and D5 dopamine receptors were used in immunoblots of bovine striatum, fresh SCO, and organ-cultured SCO, and in immunocytochemistry of the bovine, rat, and mouse SCO. Only a few tyrosine-hydroxylase fibers appeared to reach the SCO. However, virtually all the secretory ependymal and hypendymal cells of the SCO immunoreacted with antibodies against D2, D4, and D5 receptors, with the last-mentioned rendering the strongest reaction, especially at the ventricular cell pole of the secretory ependymocytes, suggesting that dopamine might reach the SCO via the cerebrospinal fluid. The antibodies against the four subtypes of receptors revealed corresponding bands in immunoblots of striatum and fresh SCO. Although the cultured SCO displayed dopamine receptors, dopamine had no apparent effect on the expression of the SCO-spondin gene/protein or on the release of RF-glycoproteins (SCO-spondin included) by SCO explants, suggesting that dopamine affects the function(s) of the SCO differently from the secretion of RF-glycoproteins.

The expression, at the mRNA level, of carotenoid biosynthetic genes from Xanthophyllomyces dendrorhous was studied by RT-PCR. The experimental conditions for the RT-PCR assay were standardized to quantify the relative transcript levels of idi, crtE, crtYB and crtI genes. This work attempted to correlate astaxanthin production with the transcript levels of carotenogenic genes in a wild-type strain (UCD 67-385) and two overproducer deregulated strains (atxS1 and atxS2). At 3 day cultures, the wild-type strain contained higher transcript levels from the crtE and crtYB genes on minimal medium than on rich medium. Similarly, carotenoid production was higher on minimal medium than on rich medium. However, carotenoid production in the atxS1 and atxS2 strains was not correlated with the transcript level of carotenogenic genes under the same experimental conditions. This result suggests that there is not a linear relationship between carotenogenic transcript levels and carotenoid biosynthesis.

Two phases may be recognized in the development of congenital hydrocephalus in the hyh mutant mouse. During embryonic life the detachment of the ventral ependyma is followed by a moderate hydrocephalus. During the first postnatal week the cerebral aqueduct becomes obliterated and a severe hydrocephalus develops. The aim of the present investigation was to elucidate the cellular phenomena occurring at the site of aqueduct obliteration and the probable participation of the subcommissural organ in this process. Electron microscopy, immunocytochemistry, and lectin histochemistry were used to investigate the aqueduct of normal and hydrocephalic hyh mice from embryonic day 14 (E-14) to postnatal day 7 (PN-7). In the normal hyh mouse, the aqueduct is an irregularly shaped cavity with 3 distinct regions (rostral, middle, and caudal) lined by various types of ependyma. In the hydrocephalic mouse, these 3 regions behave differently; the rostral end becomes stenosed, the middle third dilates, and the caudal end obliterates. The findings indicate that the following sequence of events lead to hydrocephalus: 1) denudation of the ventral ependyma (embryonic life); 2) denudation of dorsal ependyma and failure of the subcommissural organ to form Reissner fiber (first postnatal week); 3) obliteration of distal end of aqueduct; and 4) severe hydrocephalus. No evidence was obtained that NCAM is involved in the detachment of ependymal cells. The process of ependymal denudation would involve alterations of the surface sialoglycoproteins of the ependymal cells and the interaction of the latter with macrophages.

Hydrocephalic hyh mice are born with moderate hydrocephalus and a normal cerebral aqueduct. At about the fifth postnatal day the aqueduct becomes obliterated and severe hydrocephalus develops. The aim of the present investigation was to investigate the mechanism of this hydrocephalus, probably starting during fetal life when the cerebral aqueduct is still patent. By use of immunocytochemistry and scanning electron microscopy, mutant (n = 54) and normal (n = 61) hyh mouse embryos were studied at various developmental stages to trace the earliest microscopic changes occurring in the brains of embryos becoming hydrocephalic. The primary defect begins at an early developmental stage (E-12) and involves cells lining the brain cavities, which detach following a well-defined temporo-spatial pattern. This ependymal denudation mostly involves the ependyma of the basal plate derivatives. There is a relationship between ependymal denudation and ependymal differentiation evaluated by the expression of vimentin and glial fibrillary acidic protein. The ependymal cells had a normal appearance before and after detachment, suggesting that their separation from the ventricular wall might be due to abnormalities in cell adhesion molecules. The process of detachment of the ventral ependyma, clearly visualized under scanning electron microscope, is almost completed before the onset of hydrocephalus. Furthermore, this ependymal denudation does not lead to aqueductal stenosis during prenatal life. Thus, the rather massive ependymal denudation appears to be the trigger of hydrocephalus in this mutant mouse, raising the question about the mechanism responsible for this hydrocephalus. It seems likely that an uncontrolled bulk flow of brain fluid through the extended areas devoid of ependyma may be responsible for the hydrocephalus developed by the hyh mutant embryos. The defect in these embryos also includes loss of the hindbrain floor plate and a delayed in the expression of Reissner fiber glycoproteins by the subcommissural organ.

The progress on the dynamics of the radiative zone by global Doppler velocity measurements aboard SoHO (GOLF+ MDI) and with ground networks (BiSON and GONG) opens a new perspective for solar and stellar physics. It is why we prepare a new generation of solar resonant spectrometer. The objectives of the GOLF-NG instrument and its present status are described. We have demonstrated this year that most of the technical challenges have been successfully faced and the next steps are mentioned. We then recall the scientific questions that might be solved with the next generation of instruments in construction in different european laboratories to reach a complete 3D vision of our star from the core to the corona. Two formation flying missions DynaMICCS and HIRISE have been proposed to ESA in the framework of the 2015-2025 Cosmic Vision perspective to contribute to solve these questions. A strategy of measurements must be found for the next decade.

The subcommissural organ is an ependymal gland located at the entrance of the cerebral aqueduct. It secretes glycoproteins into the cerebrospinal fluid, where they aggregate to form Reissner's fiber. This fiber grows along the aqueduct, fourth ventricle, and central canal. There is evidence that the subcommissural organ is involved in the pathogenesis of congenital hydrocephalus. This organ was investigated in the mutant mouse hyh developing a congenital hydrocephalus. The central nervous system of normal and hydrocephalic hyh mice, 1 to 40 days old, was investigated using antibodies recognizing the subcommissural organ secretory glycoproteins, and by transmission and scanning electron microscopy. At birth, the affected mice displayed open communications between all ventricles, absence of a central canal in the spinal cord, ependymal denudation of the ventricles, stenosis of the rostral end of the aqueduct, and hydrocephalus of the lateral and third ventricles and of the caudal end of the aqueduct. Around the 5th postnatal day, the communication between the caudal aqueduct and fourth ventricle sealed, and hydrocephalus became severe. It is postulated that the hyh mice carry a genetic defect affecting the ependymal cell lineage. The subcommissural organ showed signs of increased secretory activity; it released to the stenosed aqueduct a material that aggregated, but it did not form a Reissner's fiber. A large area of the third ventricular wall differentiated into a secretory ependyma synthesizing a material similar to that secreted by the subcommissural organ. It is concluded that the subcommissural organ changes during hydrocephalus; whether these changes precede hydrocephalus needs to be investigated.